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  Single-Molecule Protein Phosphorylation and Dephosphorylation by Nano ore Enzymology

Harrington, L., Alexander, L. T., Knapp, S., & Bayley, H. (2019). Single-Molecule Protein Phosphorylation and Dephosphorylation by Nano ore Enzymology. ACS NANO, 13(1), 633-641. doi:10.1021/acsnano.8b07697.

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Genre: Journal Article

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 Creators:
Harrington, Leon1, Author              
Alexander, Leila T.2, Author
Knapp, Stefan2, Author
Bayley, Hagan2, Author
Affiliations:
1Schwille, Petra / Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565169              
2external, ou_persistent22              

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Free keywords: PIM KINASE INHIBITORS; PRODUCT RELEASE; DNA-MOLECULES; METAL-IONS; LABEL-FREE; BINDING; PEPTIDE; REPLICATION; ASSOCIATION; NUCLEOTIDEChemistry; Science & Technology - Other Topics; Materials Science; nanopore enzymology; phosphorylation; protein kinases; single molecule; kinase mechanism; metal ions;
 Abstract: Reversible protein phosphorylation plays a crucial and ubiquitous role in the control of almost all cellular processes. The interplay of protein kinases and phosphatases acting in opposition ensures tight dynamic control of protein phosphorylation states within the cell. Previously, engineered alpha-hemolysin pores bearing kinase substrate peptides have been developed as single-molecule stochastic sensors for protein kinases. Here, we have used these pores to observe, label-free, the phosphorylation and dephosphorylation of a single substrate molecule. Further, we investigated the effect of Mg2+ and Mn2+ upon substrate and product binding and found that Mn2+ relaxes active-site specificity toward nucleotides and enhances product binding. In doing so, we demonstrate the power and versatility of nanopore enzymology to scrutinize a critical post-translational modification.

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Language(s): eng - English
 Dates: 2019
 Publication Status: Published in print
 Pages: 9
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: ISI: 000456749900064
DOI: 10.1021/acsnano.8b07697
 Degree: -

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Title: ACS NANO
Source Genre: Journal
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Publ. Info: 1155 16TH ST, NW, WASHINGTON, DC 20036 USA : AMER CHEMICAL SOC
Pages: - Volume / Issue: 13 (1) Sequence Number: - Start / End Page: 633 - 641 Identifier: ISSN: 1936-0851